JP3698312B2 - Biosensor and biosensor manufacturing method - Google Patents

Description

[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a biosensor for quantifying a specific component in an aqueous solution and a method for producing the biosensor, and more particularly, a limited permeation of a biosensor having a selectively permeable membrane inside the reaction layer and a restricted permeable membrane outside the reaction layer. It relates to the improvement of the membrane.
[0002]
[Prior art]
In recent years, measurement using biosensors has been actively performed in the fields of medicine and analysis. Among biosensors, a current detection type glucose sensor is well known. This glucose sensor is widely used for measuring blood sugar levels and urine sugar levels.
[0003]
The most common glucose sensor uses a reaction in which glucose is converted into gluconolactone and hydrogen peroxide using glucose oxidase (GOX). That is, it reacts as shown in the following formula (1).
Glucose + O ₂ → Gluconolactone + H ₂ O ₂ (1)
(C ₆ H ₁₂ O ₆ ) GOX (C ₆ H ₁₀ O ₆ )
The reaction amount is detected as a current value by oxidizing the hydrogen peroxide generated by the equation (1) at the working electrode. The reaction at the electrode is as shown in equation (2).
H ₂ O ₂ → O ₂ + 2e ⁻ + 2H ₂ ⁺ (2)
[0004]
As is apparent from the equations (1) and (2), when O ₂ is present in excess compared to glucose, the glucose concentration in the solution is proportional to the current, so that the glucose concentration can be quantified. However, O ₂ involved in the reaction is dissolved oxygen in the sample, and the reaction is saturated at a glucose concentration (blood glucose level) of about 40 mg / dl. In practical blood glucose measurement, a range of 50 to 500 mg / dl and a urine sugar value of about 0 to 2000 mg / dl are required. Therefore, in order to widen the measurement range, 1) dilute the sample, 2) mount an electron acceptor that reacts instead of oxygen, and 3) place a permeation membrane consisting of a porous membrane on the outer layer to allow glucose to permeate. Some measures have been taken such as limiting and increasing the relative O ₂ concentration.
[0005]
Of these, 1) requires a diluting solution and a diluting device, and 2) has the problem that it can be used only once. In contrast, 3) has the advantage that the mechanism is simple and can be used repeatedly. For this reason, various researches on restricted permeable membranes have been conducted.
[0006]
One example is disclosed in Japanese Patent Laid-Open No. 9-304330. FIG. 8 shows a urine sugar measuring device using a biosensor, and FIG.
[0007]
A urine sugar measuring device 110 shown in FIG. 8 detects sugar in urine using the action of an enzyme. A sensor main body 113 is detachably attached to a measuring device main body 111 via a sensor cap 117, and the measurement is performed. Various electronic circuits are built in the instrument main body 111, and a battery or the like can be inserted through the battery cover 112. A display unit such as an LCD for displaying the measured value of urine sugar on the surface of the measuring instrument main body 111 114 is provided.
[0008]
The sensor body 113 incorporates the biosensor 100 at the tip of the sensor holder 119, and when urine is put on the biosensor 100, the sugar value in urine is automatically displayed on the display unit 114.
[0009]
As shown in FIG. 9, the biosensor 100 has a working electrode 102 and a counter electrode 103 formed on an insulating substrate 101, and a selective permeable membrane 104, an immobilized enzyme membrane 105, and a polycarbonate porous membrane 108 are sequentially laminated thereon. Yes. Here, the polycarbonate porous membrane acts as the limiting permeable membrane 108, and the measurement range of blood glucose level and the like is expanded to 0 to 500 mg / dl.
[0010]
[Problems to be solved by the invention]
However, the cited conventional method has the following problems. First, the film used as the limiting permeable membrane 108 is expensive. Since this film is produced such that a polycarbonate film is irradiated with a neutron beam and the altered portion altered by the neutron beam is chemically etched to form a predetermined through hole, it cannot be made as cheap as an ordinary resin material.
The second problem is that a process of attaching a film to each sensor is required. Such an operation requires a larger number of man-hours than a normal wafer film forming process. In particular, it is difficult to select an adhesive, which also increases the cost.
A third problem is that film parameters such as the hole diameter and aperture ratio cannot be easily changed. Film production requires large-scale equipment, making it difficult to produce a variety of products in small quantities.
[0011]
The present invention has been made to solve the above-mentioned problems. The problem to be solved by the present invention is that the opening ratio of the limiting permeable membrane used in the biosensor can be easily set by a simple process, and the glucose concentration is 0. A biosensor that does not saturate to about 5000 mg / dl and a method for manufacturing the biosensor are provided.
[0012]
[Means for Solving the Problems]
The invention according to claim 1 is a biosensor 10 having a selectively permeable membrane 4 below the reaction layer (immobilized enzyme membrane) 5 and a restricted permeable membrane 8 outside the reaction layer 5, which is a restricted permeable membrane. A plurality of minute projections 6 provided on a base layer 8 of a photosensitive material using a non-organic solvent as a peeling solution, and an impermeable film 7 provided around the plurality of projections 6. The biosensor is characterized in that the plurality of protrusions 6 are removed to leave a water-impermeable film, thereby forming a limited permeable membrane 8 having a plurality of small openings.
[0013]
The invention according to claim 2 is the biosensor according to claim 1, wherein the biosensor 10 is a current detection type biosensor.
[0014]
The invention according to claim 3 is the biosensor according to claim 1, wherein the biosensor 10 is a glucose sensor.
[0015]
The invention according to claim 4 is the biosensor according to claim 1, wherein the biosensor 10 is a glucose sensor comprising a hydrogen peroxide electrode and glucose oxidase.
[0016]
The invention according to claim 5 is the biosensor according to claim 1, wherein the biosensor 10 is a biosensor using an ion-sensitive field effect transistor as a transducer.
[0017]
The invention according to claim 6 is a method of manufacturing a biosensor having a permselective membrane below the reaction layer and a restrictive permeation membrane outside the reaction layer, wherein the organic layer is formed on the underlayer of the restrictive permeation membrane. The method comprises a step of forming a plurality of minute protrusions using a photosensitive material having a solvent as a peeling solution, a step of forming a water-impermeable film around the protrusions, and a step of removing the protrusions. This is a method for manufacturing a biosensor.
[0018]
The invention according to claim 7 is the method for producing a biosensor according to claim 6, wherein the stripping solution of the photosensitive material is an aqueous sodium hydroxide solution.
[0019]
The invention according to claim 8 is the biosensor manufacturing method according to claim 6, wherein the water-impermeable film is a resin curable at 80 ° C. or lower.
[0020]
The invention according to claim 9 is the biosensor manufacturing method according to claim 6, characterized in that the water-impermeable film is mainly composed of any one of silicone, polyurethane, polyacrylic acid, and fluororesin. .
[0021]
The invention according to claim 10 is the method for producing a biosensor according to claim 6, wherein the protrusion 6 is cylindrical or conical.
[0022]
The invention according to claim 11 is characterized in that, in the step of exposing the photosensitive material, overexposure or underexposure is performed so that the protrusion 6 is made thinner than when exposed under normal conditions. This is a method for manufacturing a biosensor.
[0023]
According to a twelfth aspect of the present invention, there is provided the biosensor manufacturing method according to the sixth aspect, wherein the diameter of the protrusion 6 is 1 μm or less.
[0024]
A thirteenth aspect of the present invention is the biosensor manufacturing method according to the sixth aspect of the present invention, wherein the aperture ratio of the restricted permeable membrane 8 is 0.01 to 10%.
[0025]
The invention according to claim 14 is the method of manufacturing a biosensor according to claim 6, wherein a plurality of biosensors 10 are simultaneously produced on the same substrate 14.
[0026]
DETAILED DESCRIPTION OF THE INVENTION
Hereinafter, the biosensor and the biosensor manufacturing method of the present invention will be described in detail with reference to FIGS.
[0027]
FIG. 1A is a plan view of a biosensor showing an embodiment of the present invention, and FIG. 1B is a cross-sectional view taken along the line AA ′ in FIG. 1A and 1B, a substantially rectangular working electrode 2 and a counter electrode 3 are formed on an insulating substrate 1 and connected to substantially rectangular electrode pads 12A and 12B. Next, the permselective membrane 4 is laminated. The permselective membrane 4 is installed for the purpose of eliminating molecules having a large molecular weight only through the H ₂ O _{2 to be} detected. Above this, an immobilized enzyme membrane 5 is formed. A projection 6 made of a photoresist using a non-organic solvent as a stripping solution is formed thereon.
[0028]
The reason for using a photoresist with a non-organic solvent as a stripping solution is as follows. As the permselective membrane 4, cellulose, ion exchange resin or the like is often used. These materials are generally soluble in organic solvents. Therefore, when a photoresist using an organic solvent as a stripping solution is used, the permselective film 4 is dissolved during stripping. For this reason, a photoresist using a non-organic solvent as a stripping solution must be used. A material that satisfies this requirement includes a solder resist used for a printed wiring board, and can be obtained at a low price.
[0029]
A non-permeable membrane 7 is formed around the protrusion 6 of the selectively permeable membrane 4. The water-impermeable film 7 is preferably one that can be cured at room temperature. This is because many of the enzymes used in biosensors are deactivated at high temperatures and lose their catalytic function. Even in GOX, which is relatively resistant to heat, the heat-resistant temperature is about 80 ° C. Suitable resins for the water-impermeable film 7 include silicone, polyurethane, polyacrylic acid, fluororesin and the like. After the water-impermeable membrane 7 is cured, the protrusion 6 is removed with a stripping solution, and the biosensor 10 having the limited permeable membrane 8 is completed.
[0030]
Although FIG. 1B shows a cross-sectional view of the biosensor 10 from which the single protrusion 6 is removed, actually, the plan view of FIG. 3A and the BB ′ cross-sectional arrow of FIG. As shown in FIG. 3 (B), a selective permeable membrane 4 and a plurality of biosensor elements comprising a working electrode 2 and a counter electrode 3 and electrode pads 12A and 12B on an insulating substrate 14 such as a large area glass are provided. A plurality of biosensor elements are integrally formed in a state where the immobilized enzyme membrane 5 and the impermeable membrane 7 are sequentially formed, and then a single biosensor 10 is cut into a pentagonal shape, A single biosensor 10 as shown in (B) is obtained.
[0031]
2A to 2F are schematic cross-sectional views showing the steps of the biosensor 10 shown in FIGS. 1A and 1B. Hereinafter, it demonstrates according to the order of a process. In FIG. 2A, a conductor film is formed on the insulating substrate 1 by vapor deposition or the like, and the working electrode 2 and the counter electrode 3 are formed by photolithography. In FIG. 2B, the selective transmission film 4 is formed on the entire surface by spin coating or the like. In FIG. 2C, the immobilized enzyme film 5 is formed on the entire surface by spin coating or the like, and a photoresist 9 using a non-organic solvent as a peeling solution is formed on the entire surface of the immobilized enzyme film 5, and then FIG. ), A large number of minute protrusions 6 made of a photoresist 9 are formed by exposure and development. In FIG. 2E, a water-impermeable resin solution is applied by spin coating or the like and dried to form a water-impermeable film 7 around the protrusions 6. In FIG. 2 (F), the protrusion 6 is removed using a stripping solution. In this way, the limited permeable membrane 8 having the minute permeable holes 11 and made of an impermeable material is formed.
[0032]
An embodiment of the current detection type biosensor shown in the above-described embodiments of FIGS. 1 to 3 will be described below.
[0033]
[Example 1]
A glass plate is used as the insulating substrate 14 shown in FIGS. 3A and 3B, the size is 50 mm × 60 mm, platinum (Pt) is vapor-deposited on the glass plate, and the action is performed by a lift-off method, milling, or the like. 100 sets of biosensor elements in which the electrode 2, the counter electrode 3, and the electrode pads 12 </ b> A and 12 </ b> B are patterned are formed. The conductive film may be gold (Au) or silver (Ag).
[0034]
Next, an alcohol solution of an ion exchange resin {trademark, Nafion} is applied to the entire surface by spin coating and dried at room temperature to obtain a selective oxide film 4.
[0035]
Next, a mixed solution of bovine serum albumin, glucose oxidase, and glutaraldehyde is applied to the entire surface and dried at room temperature to form an immobilized enzyme film 5.
[0036]
Next, a photoresist 9 whose stripping solution is a non-organic solvent is applied as shown in FIG. 2C, and this photoresist 9 is exposed and developed to form a protrusion 9 as shown in FIG. .
[0037]
As a mask pattern for exposing and developing the photoresist 9, one circle having a diameter of 1 μm was placed on the entire surface at a density of 10 ⁸ / cm ² in 10 μm square. A 1% sodium carbonate (Na ₂ CO ₃ ) aqueous solution was used as the developer. Next, a silicone resin solution was applied to the entire surface with a thickness of 1 μm by spin coating. Next, the protrusions 6 were removed using a 4% aqueous sodium hydroxide (NaOH) solution. In this way, the limited permeable membrane 8 having many through holes 6 having a diameter of 1 μm was formed. In photolithography using ultraviolet rays, the limit of patterning is about 0.1 μm, and it is relatively easy to make a pattern of 1 μm. The aperture ratio of the limited permeable membrane 8 obtained in this way is a little less than 1%. Since the measurement range of urine sugar and the like is inversely proportional to the aperture ratio, in this case, the measurement range is designed to be about 100 times the original. Next, the insulating substrate 14 was divided to obtain 100 glucose sensors 10.
[0038]
The glucose sensor 10 was evaluated using a potentiostat (constant voltage stabilized power supply). In the evaluation using the phosphate buffer containing glucose, a linear calibration curve was obtained in the glucose concentration range of 0 to 5000 mg / dl. Next, when serum containing 500 mg / dl glucose was repeatedly measured 10 times, the coefficient of variation (standard deviation / average value) was 0.3%. Next, urine containing 2000 mg / dl glucose was measured in the same manner. As a result, the coefficient of variation was 10%, and the magnitude of the output current decreased by 10% after 10 measurements.
[0039]
In Example 1, a silicone resin is used as the water-impermeable film 7, but any resin that can be cured at 80 ° C. or lower can be used. For example, a polyurethane resin, a polyacrylic acid resin, a fluorine resin, or the like can be used.
[0040]
Further, the aperture ratio of the through holes 11 formed on the restricted permeable membrane 8 will be described with reference to FIGS. 4 (A) and 4 (B). FIG. 4A schematically shows the through-hole 11 on the working electrode 2 formed in the insulating substrate 1 used in the biosensor 10 of another embodiment to be described later, but in principle, on the working electrode 2. What is necessary is just to make it control the aperture ratio. Here, the aperture ratio indicates the ratio of the total area of the through holes to the circular area of the working electrode. FIG. 4B shows a saturation characteristic curve when the vertical axis represents the output current obtained between the working electrode 2 and the counter electrode 3 of the current detection type biosensor, and the horizontal axis 7 represents the glucose concentration. The saturation characteristic curve 22 in FIG. 4B is a saturation characteristic curve when the restricted permeable membrane 8 is not provided, and 23 is a saturation characteristic curve when the polycarbonate porous membrane shown in FIG. 9 is used as the restricted permeable membrane 108. In the first embodiment of the present invention, the case where the aperture ratio is selected to be 1% has been described. However, the diameter of the through hole 11 of the restricted transmission film 8 and the aperture ratio are designed appropriately for the mask pattern used during exposure and development of the photoresist 9. When the diameter is 1 μm or less (excluding 0) and the aperture ratio is selected from 0.01 to 10%, the saturation point of the glucose concentration is set to 50 to 500 mg / dl at the time of blood glucose measurement, and urine sugar A characteristic saturation curve 25 having a linear calibration curve can be easily and freely obtained from 0 to 2000 mg / dl (characteristic saturation curve 24) at the time of value measurement or a glucose concentration higher than 5000 mg / dl.
[0041]
FIGS. 5A and 5B show another embodiment of the present invention. The difference from FIGS. 1A and 1B is that Pt, Ag, Au, etc. are patterned on the insulating substrate 1. 1A, a semicircular counter electrode 3 and a reference electrode (reference electrode) 13 and electrode pads 12A, 12B, and 12C are provided so as to surround the substantially circular (2 mmφ) working electrode 2 from the left and right. In the counter electrode 3 of FIG. 5B, the reference electrode 13 is also used as the counter electrode 3, but in FIGS. 5A and 5B, an independent reference electrode 13 is provided to improve the measurement accuracy.
[0042]
The selective permeable membrane 4 coated on the working electrode 2, the counter electrode 3, and the reference electrode 13, the immobilized enzyme membrane 5, the impermeable membrane 7 and the restricted permeable membrane 8 comprising the through holes 11 are shown in FIGS. ).
[0043]
In FIGS. 1 (A) and 1 (B), the amount of O ₂ consumed by the reaction of glucose as a substrate with the enzyme of GOX is measured by current conversion according to the reaction formulas (1) and (2). However, it is also possible to use an ion sensitive transistor (ISFET) or the like, which will be described later, and it is also possible to use alcohol dehydrogenase other than glucose, penicillinase or the like as the enzyme. That is, the biosensor and biosensor manufacturing method of the present invention can also be applied to biosensors other than glucose sensors.
[0044]
Further, another embodiment of the biosensor and the biosensor manufacturing method of the present invention will be described with reference to FIGS.
[0045]
6 (A) and 6 (B), the configuration is the same as that of the biosensor of FIGS. 1 and 2 except for the formation of the protrusion 6. After the immobilized enzyme film 5 is formed, a photoresist 9 using a non-organic solvent as a stripping solution is applied to the entire surface. At this time, it is desirable to make the resist thickness thinner than in the case of FIG. Next, as shown in FIG. 6A, the resist 9 is exposed from the light source 14 using a predetermined mask 16. When using a negative resist, the exposure amount at this time is reduced by several tens of percent from the normal condition. Next, a protrusion 6 obtained by developing the photoresist 9 is formed. By doing in this way, the projection part 6 of the diameter narrower than the pattern 17 of the mask 16 is obtained.
[0046]
In FIG. 6 (A), if the diameter of the pattern 17 on the mask 16 is 1 μm, a cylindrical protrusion 6 having −Δl of 1 μm is formed.
[0047]
Conversely, when a positive resist is used as shown in FIG. 6B, the same result can be obtained by overexposure. That is, if the diameter of the pattern 17 of the transmissive part on the mask 16 is 1 μm, the overexposure results in the projection 6 having a diameter smaller than the diameter of the transmissive part. In the case of FIG. 6B, the protrusion 6 is formed in a conical shape.
[0048]
[Example 2]
In the same manner as in Example 1, up to the immobilized enzyme membrane 5 was formed. Next, a negative photoresist 9 was applied to a thickness of 0.5 μm. The mask pattern was the same as in Example 1. Next, the photoresist 9 was exposed by reducing the exposure amount by 20% from that in Example 1. When this was developed, a projection 6 having a diameter of 0.7 μm was obtained. Next, the silicone solution was spin-coated as a water-impermeable film 7 at a thickness of 0.5 μm and dried at room temperature. Next, the protrusion 6 was removed using a 4% NaOH aqueous solution.
[0049]
This glucose sensor 10 was evaluated using a potentiostat. In the evaluation using the phosphate buffer containing glucose, a linear calibration curve was obtained in the glucose concentration range of 0 to 8000 mg / dl. Next, when serum containing 500 mg / dl glucose was repeatedly measured 10 times, the coefficient of variation was 0.3%. Next, urine containing 2000 mg / dl glucose was measured in the same manner. The coefficient of variation was 5%, and no systematic decrease in output current was observed. The reason why the reproducibility was better than that of Example 1 was that contaminants (proteins and the like) infiltrated into the immobilized enzyme membrane 5 as the pore diameter of the through-hole 11 in the restricted permeable membrane 8 became smaller. It seems that it was prevented. It can be seen that it is better to change the mask and further reduce the hole diameter of the through hole 11 in order to increase the accuracy.
[0050]
FIG. 7 is a schematic side sectional view of a biosensor 10A in which an insulating substrate 1 portion is an ISFET (Ion Sensitive FET) as a chemical sensor to which the manufacturing method of the present invention can be applied. ) Restricted permeation in which a permeation hole 11 is formed in the permselective membrane 4, the immobilized enzyme membrane 5, and the water-impermeable membrane 7 instead of the gate provided between the source 18 and the drain 19 on the insulating substrate 20 such as silicon 21. The film 8 is formed sequentially.
[0051]
Unlike the current detection type biosensor 10, the above-described ISFET detects an electrochemical potential change using the FET 21. When a sample (aqueous solution) is immersed in the biosensor 10, an ion-sensitive layer such as the immobilized enzyme membrane 5. Since the potential of the surface of the FET 21 changes via the voltage, the amount of change in the potential is proportional to the PH value in the aqueous solution, and the hydrogen ion concentration (PH) is measured.
[0052]
In this case, the limited permeable membrane 8 is also a biosensor capable of easily controlling the aperture ratio of the through holes 11 formed in the impermeable membrane 7 by photolithography using an inexpensive photosensitive material, and a method of manufacturing the biosensor. can get.
[0053]
【The invention's effect】
As described above, according to the present invention, a biosensor having a wide measurement range can be obtained easily and inexpensively. In addition, since the pore size and aperture ratio of the restricted permeable membrane can be freely designed, it is possible to manufacture a biosensor and a biosensor having arbitrary characteristics without increasing costs.
[Brief description of the drawings]
1A and 1B are a plan view and a cross-sectional schematic view showing an embodiment of the present invention.
FIG. 2 is a schematic cross-sectional view showing an outline of the process of FIG. 1;
FIGS. 3A and 3B are a plan view and a schematic cross-sectional view showing a configuration when obtaining the biosensor of the present invention at the same time. FIGS.
FIG. 4 is an explanatory diagram of an aperture ratio on the working electrode of a biosensor used in the present invention, and a saturation characteristic curve explanatory diagram showing a relationship between a glucose concentration of the biosensor and a detected output current.
FIGS. 5A and 5B are a front view and a cross-sectional schematic view showing another embodiment of the present invention. FIGS.
FIG. 6 is a sectional side view of a main part showing a method for producing a restricted permeable membrane of a biosensor of the present invention.
FIG. 7 is a schematic sectional view showing another embodiment of the biosensor of the present invention.
FIG. 8 is an external view of a conventional urine sugar measuring device.
FIG. 9 is a schematic cross-sectional view showing an example of a conventional biosensor.
[Explanation of symbols]
1,14,20 ... Insulating substrate, 2,102 ... Working electrode, 3,103 ... Counter electrode, 4,104 ... Permselective membrane, 5,105 ... Immobilized enzyme membrane, 6 ... Projection 7, impervious film, 8 restricting permeable film, 9 photoresist, 11 through hole, 12 A, 12 B, 12 C electrode pad, 13 reference electrode, 21 FET, 108 Polycarbonate porous membrane

Claims (14)

A biosensor having a permselective membrane below the reaction layer and having a restricted permeation membrane outside the reaction layer,
A plurality of minute protrusions provided by a photosensitive material having a non-organic solvent as a stripping solution on the underlayer of the limiting permeable membrane;
An impermeable film provided around the plurality of protrusions,
A biosensor comprising the restricted permeation membrane having a plurality of small openings leaving the plurality of protrusions to leave the impermeable membrane. The biosensor according to claim 1, wherein the biosensor is a current detection type biosensor. The biosensor according to claim 1, wherein the biosensor is a glucose sensor. The biosensor according to claim 1, wherein the biosensor is a glucose sensor comprising a hydrogen peroxide electrode and glucose oxidase. The biosensor according to claim 1, wherein the biosensor is a biosensor using an ion-sensitive field effect transistor as a transducer. A method for producing a biosensor having a permselective membrane below the reaction layer and having a restricted permeation membrane outside the reaction layer,
Forming a plurality of minute protrusions using a photosensitive material having a non-organic solvent as a stripping solution on the underlayer of the limiting permeable membrane;
Forming a water-impermeable film around the protrusion, and
A method for producing a biosensor comprising the step of removing the protrusion. The biosensor manufacturing method according to claim 6, wherein the stripping solution of the photosensitive material is an aqueous sodium hydroxide solution. The biosensor manufacturing method according to claim 6, wherein the water-impermeable film is a resin curable at 80 ° C. or lower. 7. The method of manufacturing a biosensor according to claim 6, wherein the water-impermeable film contains silicone, polyurethane, polyacrylic acid, or a fluororesin as a main component. The method of manufacturing a biosensor according to claim 6, wherein the protrusion is cylindrical or conical. 7. The method of manufacturing a biosensor according to claim 6, wherein in the step of exposing the photosensitive material, overexposure or underexposure is performed to make the protrusions thinner than when exposed under normal conditions. . The biosensor manufacturing method according to claim 6, wherein a diameter of the protrusion is 1 μm or less. The method for producing a biosensor according to claim 6, wherein an opening ratio of the restricted permeable membrane is 0.01 to 10%. The biosensor manufacturing method according to claim 6, wherein a plurality of the biosensors are simultaneously manufactured on the same substrate.

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